Fungal pathogens represent one of the most serious threats to agriculture production. The development of crop varieties that simultaneously control different pathogens is an attracting strategy to increase host
plant resistance. This can be obtained by improving the pre-existing plant defence mechanisms such as the structure and composition of the plant cell wall, which is one of the first barriers encountered by the
microbial pathogens during plant tissue colonization. To overcome this obstacle, most fungal pathogens produce a variety of enzymes that degrade the wall polysaccharides; among them, pectin degrading
enzymes are among the first to be secreted by the pathogens to enter and spread into the plant tissue.
Pectin in the plant cell wall is secreted in a highly methylesterified form and is demethylesterified in muro by pectin methylesterase (PME). The activity of PME is regulated by specific protein inhibitors (PMEIs).
Since highly methylesterified pectin can be less susceptible to hydrolysis by pectic enzymes such as fungal endopolygalacturonases (endo-PGs), the inhibition of endogenous PME by PMEI might increase pectin resistance to degradation by fungal PGs. In order to verify the possibility to improve wheat resistance by modifying the degree of pectin methylesterification, a number of wheat Triticum durum cv.
Svevo lines expressing the pectin methylesterase inhibitors either AcPMEI (from Actinidia chinensis) or AtPMEI-1 (from Arabidopsis thaliana) has been produced. Both AcPMEI and AtPMEI-1 endow wheat with a stronger inhibitory activity of endogenous PME. Only one line expressing AtPMEI-1 shows
reduced endogenous PME activity while 12 lines expressing AcPMEI show different levels of reduced PME activity. Three Acpmei lines were further characterized on the level of expression and the activity of the transgenic protein. AcPMEI was found to be correctly targeted to the apoplast. The two Acpmei lines expressing high level of the inhibitor, MJ15-151 and MJ15-69, showed no obvious phenotypic differences with the control plants but a significant increase of the degree of methylesterification (DM) of
about 20% and 32%, and a different methylester distribution compared to the wild-type plants, respectively. This increased DM and modified methyl distribution of pectin was correlated with a significant reduction of disease symptoms caused by the foliar fungal pathogen Bipolaris sorokiniana or the floral pathogen Fusarium graminearum.
To further protect the cell wall pectic component, a pyramiding approach has been undertaken to combine
PMEI and the polygalacturonase-inhibiting protein (PGIP). The cross between the Acpmei lines and a new developed durum wheat transgenic line expressing Pvpgip2 has been performed and the F1 progeny analyzed for the presence of both transgenes.
Finally, to select additional fungal pathogens to be tested in the cross Acpmei/Pvpgip2 and in the parental lines, infection experiments with the biotrophic fungal pathogen Claviceps purpurea were performed on a stable transgenic bread wheat line expressing Pvpgip2 and a slight, even significant, reduction of symptoms in the transgenic line compared to the control plants was observed....more